Project Summary The overall goal of this project is to increase our understanding of neurobiological signaling process that mediates physiological and behavioral effects of dopamine. This proposal focuses on a novel regulation of dopamine D2 receptor (D2R) signaling in the midbrain by RGS2 (regulator of G protein signaling 2) protein. Dysfunctional midbrain D2R is implicated in neurological and psychiatric diseases. However, little knowledge is known about the mechanisms of midbrain D2R signaling. This proposal will delineate the mechanistic actions of RGS2 on D2R signaling and interrogate RGS2 in dopaminergic neurons as a critical modulator of midbrain D2R-mediated dopamine physiology and behavior. D2R signals via its coupled G?i/o protein to mediate cellular and behavioral responses to stimuli. The family of RGS proteins is a key negative modulator of D2R signaling by accelerating GTP hydrolysis and terminating G protein signaling. To date, no study has examined the associations between specific RGS proteins and D2R signaling in midbrain dopaminergic neurons. We find that amphetamine self-administration increases RGS2 protein levels and decreases D2R-stimulated G protein activation in rat midbrain. Moreover, RGS2 and somatodendritic D2R are both expressed in midbrain dopaminergic neurons. Thus, RGS2 and D2R may be functionally linked which has never been examined in a neuronal system. Using neuroblastoma N2A cells as a model system, we made the novel observation that RGS2 negatively regulates D2R-mediated G?i/o signaling. Moreover, RGS2 couples with D2R via its N-terminus. Thus, we hypothesize that RGS2 directly interacts with D2R to engage a unique G?i/o signaling pathway that controls physiological and behavioral responses of D2R in midbrain dopaminergic neurons. This hypothesis will be tested in two specific aims: 1) assess whether RGS2 directly interacts with D2R to control D2R-mediated G protein signaling in neuron-like cell lines; and 2) determine the physiological and behavioral significance of the RGS2-D2R interaction in dopaminergic neurons of ventral tegmental area. This proposal may expand the repertoire and diversity of D2R regulation and has a potential to identify RGS2 as a novel component of the D2R signaling network in vivo